We would like to thank Drs. Stratmann and Alvi for their interest in our study.1In the study, we tried to determine the comparative neurotoxic effects of the volatile anesthetics desflurane, isoflurane, and sevoflurane on the developing mouse brain. Accordingly, we first established comparative anesthetic potencies for the three volatile agents and then found neuroapoptosis in neocortical neurons to be similar when animals were exposed to equipotent concentrations of 0.6 minimum alveolar concentration (MAC) for 6 h. Stratmann and Alvi now raise two important issues: first, how are the neurotoxic effects of the different anesthetic agents best compared in small animal model systems, and second, how does one ensure similar anesthetic potency throughout the exposure.
For their first comment, Stratmann and Alvi point out that several studies have observed differential effects of sevoflurane on neuronal cells compared with isoflurane or propofol.2,–,5However, none of the studies comparing isoflurane with other anesthetics have established a similar degree of anesthetic potency among the experimental groups in their particular model systems2,–,6; instead, they have used concentrations extrapolated from adult studies. Moreover, three of the quoted studies used in vitro models with dissociated cell cultures,2,–,4which may not represent an adequate model system to quantify the degree of neurotoxicity observed in live animals. Accordingly, several in vitro studies have been unable to detect any neurotoxic anesthetic effects, even when using isoflurane.7,8Although the mechanism of anesthetic neurotoxicity is just beginning to be understood,9,10it has been suggested to be associated with anesthetic interference with neuronal networks, which cannot be modeled well in dissociated neuronal cultures.11
Another important question in comparing different studies of anesthetic neurotoxicity is how brain cell death was quantified. Neuronal cell death after anesthetic exposure is not homogeneously distributed throughout the brain but varies by brain region and even within a particular brain region, and may differ depending on which animal species or anesthetic agents are studied. Accordingly, we selected a region that in our model repeatedly has shown significant anesthesia-induced neuroapoptosis (i.e. , layer II/III of the neocortex; see figure 3 from the original article). An observer unaware of group assignment repeatedly was able to distinguish sections from previously anesthetized animals from those of unanesthetized littermates but was not able to differentiate among the three anesthetic regimens. Two quantitative methods, cell counts of apoptotic neurons and a colorimetric assay of the activity of the apoptotic enzyme caspase 3, could not be differentiated among the three anesthetic regimens. Thus, we are confident in our findings that a 6-h exposure to 7.4% desflurane, 1.5% isoflurane, or 2.9% sevoflurane, respectively, produced similar degrees of neuroapoptosis in the superficial neocortex of newborn CD1/C57BL/6J hybrid mice. Our qualitative assessment makes significant differences in neuroapoptosis in other brain regions unlikely; however, we cannot rule out that quantitative cell counts may result in differences and that comparative results in other species may differ.
The second point raised by Stratmann and Alvi relates to the important issue of establishing and maintaining equivalence between different anesthetic agents when comparing their neurotoxic potency. To accomplish this, one can either determine the true MAC value for each anesthetic while ensuring stable environmental factors12(as we did in our study) or use the approach taken by Stratmann et al. and repeatedly measure MAC in a cohort of subjects while environmental factors change dramatically.13In the former approach, MAC has been found to remain constant during anesthetic exposure,14whereas in the latter approach it is not surprising that MAC will change over time because of the changes in environmental factors, such as alterations in sodium concentrations, temperature, oxygenation, carbon dioxide concentrations, glucose, acid base status, or hemoglobin concentrations, which all can lead to significant changes in MAC.15,16Moreover, MAC differs by age, species, and even between strains of the same species.17This would suggest that Stratmann and Alvi's extensive comparisons of their data obtained in rats may have only limited applicability to our study in mice. Contrary to the substantial hypercapnia and acidosis observed by Stratmann et al. in their rat studies,13,18acid–base derangements were far less severe in our study and did not differ among the desflurane, isoflurane, and sevoflurane groups, suggesting that any effects on MAC would be expected to be less dramatic and similar among the three anesthetic groups.
However, Stratmann and Alvi's comment raises another interesting question: whether the absolute concentration of a drug, which differed by a factor of almost 5 between isoflurane and desflurane, or its MAC concentration determines its neurotoxic effects. Our finding of similar toxicity using equipotent concentrations of the three anesthetic agents seems to suggest that anesthetic potency is the determining factor. If these findings in animals applied to the clinical setting as well, where anesthetics are dosed according to their potency to provide amnesia and immobility during surgery, no benefit would be gained by choosing one anesthetic agent over another.
George K. Istaphanous, M.D., John J. McAuliffe, M.D., M.B.A., Andreas W. Loepke, M.D., Ph.D.
**Cincinnati Children's Hospital Medical Center, University of Cincinnati College of Medicine, Cincinnati, Ohio. firstname.lastname@example.org